Ebola (EBOV) and Marburg (MARV) viruses are NIAID Category A pathogens and cause severe hemorrhagic fevers in humans and nonhuman primates with mortalities as high as 90%. There are currently no licensed vaccines for preventing disease caused by these viruses. Recent successes in eliciting protective immunity in nonhuman primates with recombinant vesicular stomatitis virus (VSV) or adenovirus- vectored EBOV or MARV vaccines suggest that effective vaccines can be developed. Despite the promising results, concerns have been raised about the use of these vaccine platforms in humans. The recombinant adenovirus vaccines are replication deficient;thus, very high doses are needed to induce protective immunity. More importantly, because adenoviruses are common human pathogens, most humans have pre- existing immunity, which greatly interferes with the ability of the vector to induce an immune response to the recombinant gene product. In addition to naturally occurring preexisting immunity, the broad use of recombinant adenovirus vectors complicates the field of vaccine development because vector antibodies induced as the result of vaccination would suppress the responses to an independent vaccine utilizing a similar delivery strategy. In contrast, the VSV vector is a live virus vaccine that does not commonly infect humans;however, it is a human pathogen and significant safety concerns about its use have been raised. By comparison, DNA vaccines have been proven safe in many human clinical trials and there is no possibility of vector immunity. While DNA vaccines have lacked sufficient potency in humans, the development of in vivo electroporation delivery systems has the potential to address this shortcoming. The requested funding will not only facilitate progress toward safe and effective combination vaccines for Ebola (EBOV) and Marburg (MARV) viruses, but will also advance the development of a vaccine delivery platform that has the capacity to address many of the performance attributes desired in the field of biodefense, namely a readily manufactured, stable vaccine with sufficient potency to provide protective efficacy against multiple pathogens after few immunizations. As such, the research proposed directly addresses the goals of this solicitation. Specific aims of the research are to: (1) Generate improved DNA vaccine constructs for two species of EBOV and two strains of MARV through codon-optimization of the GP genes of each virus;(2) Compare the immunogenicity of the individual DNA vaccine candidates when delivered to mice in various dosages using electroporation via intramuscular or intradermal routes;(3) Evaluate and compare the protective efficacy of combined vs. individual DNA vaccines in mice after immunization by electroporation;(4) Demonstrate the immunogenicity and protective efficacy of combined EBOV and MARV DNA vaccines in non-human primates.